Organotypic slice cultures for analysis of proliferation, cell death, and migration in the embryonic neocortex.

ABSTRACT

Dynamic cellular interactions during neocortical neurogenesis are critical for proper cortical development, providing both trophic and tropic support. Although cell proliferation and programmed cell death have been characterized in dissociated primary cell cultures, many in vivo processes during cortical neurogenesis depend on cell-cell interactions and therefore on the three-dimensional environment of the proliferating neuroblasts and their progeny. Here we describe a murine organotypic neocortical slice preparation that retains major morphological and functional in vivo characteristics of the developing neocortex and is viable (exhibits very low levels of cell death) for up to three days. Moreover, this slice preparation is amenable to direct experimental manipulation of potential diffusible regulators of neurogenesis. Using a variety of biochemical and physiological methods including time-lapse and quantitative confocal microscopy, we demonstrate that this system can be used effectively to investigate cellular mechanisms important for brain growth and maturation, including neurogenesis, apoptosis, and neuronal migration.